DataSheet_1_Zinc acquisition and its contribution to Klebsiella pneumoniae virulence.docx

Klebsiella pneumoniae is a World Health Organization priority pathogen and a significant clinical concern for infections of the respiratory and urinary tracts due to widespread and increasing resistance to antimicrobials. In the absence of a vaccine, there is an urgent need to identify novel targets for therapeutic development. Bacterial pathogens, including K. pneumoniae, require the d-block metal ion zinc as an essential micronutrient, which serves as a cofactor for ~6% of the proteome. During infection, zinc acquisition necessitates the use of high affinity uptake systems to overcome niche-specific zinc limitation and host-mediated nutritional immunity. Here, we report the identification of ZnuCBA and ZniCBA, two ATP-binding cassette permeases that are highly conserved in Klebsiella species and contribute to K. pneumoniae AJ218 zinc homeostasis, and the high-resolution structure of the zinc-recruiting solute-binding protein ZniA. The Znu and Zni permeases appear functionally redundant with abrogation of both systems required to reduce K. pneumoniae zinc accumulation. Disruption of both systems also exerted pleiotropic effects on the homeostasis of other d-block elements. Zinc limitation perturbed K. pneumoniae cell morphology and compromised resistance to stressors, such as salt and oxidative stress. The mutant strain lacking both systems showed significantly impaired virulence in acute lung infection models, highlighting the necessity of zinc acquisition in the virulence and pathogenicity of K. pneumoniae.

肺炎克雷伯菌（Klebsiella pneumoniae）是世界卫生组织（World Health Organization, WHO）划定的优先病原体，由于其对各类抗菌药物（antimicrobials）的耐药性呈广泛且上升趋势，已成为引发呼吸道与尿路感染的重大临床关注点。目前尚无针对该菌的获批疫苗，因此亟需鉴定新型治疗药物开发靶点。包括肺炎克雷伯菌在内的细菌病原体，均需以d区金属离子锌（d-block metal ion zinc）作为必需微量营养元素，锌可作为约6%蛋白质组（proteome）的辅因子发挥功能。在感染过程中，细菌获取锌需借助高亲和力摄取系统，以克服定植微环境中的锌限制与宿主介导的营养免疫。本研究成功鉴定出ZnuCBA与ZniCBA两种ATP结合盒转运蛋白（ATP-binding cassette permeases），二者在克雷伯菌属物种中高度保守，可参与肺炎克雷伯菌AJ218的锌稳态调控，同时解析了锌募集溶质结合蛋白ZniA的高分辨率结构。Znu与Zni转运系统在功能上似乎存在冗余，仅同时敲除这两套系统才能降低肺炎克雷伯菌的锌积累水平。同时敲除两套系统还会对其他d区金属元素的稳态产生多效性影响。锌限制会扰动肺炎克雷伯菌的细胞形态，并削弱其对盐胁迫与氧化应激等逆境的抗性。同时缺失两套系统的突变菌株在急性肺感染模型中的毒力显著受损，这凸显了锌摄取能力对于肺炎克雷伯菌的致病力与致病性的关键必要性。